The effect of maternal obesity on fatty acid transporter expression and lipid metabolism in the full‐term placenta of lean breed swine

This study was conducted to evaluate the influence of back‐fat thickness (BF), at mating of sows, on the maternal and newborn circulating lipids, expression of placental fatty acids (FA) transporters and lipid accumulation in placenta. Full‐term placentas were obtained by vaginal delivery from BFI (9–14 mm; n = 37), BFII (15–19 mm; n = 43) and BFIII (20–27 mm; n = 38) sows according to BF at mating, and frozen placental sections were analysed for fat accumulation. Blood samples were collected from the sows of day 105 pregnancy and from cord blood at delivery. mRNA and protein expression levels were evaluated with real‐time RT‐PCR and Western blotting. Our results demonstrated that BFII females had significantly increased litter weight and placental efficiency, decreased maternal triglyceride (TG) and non‐esterified fatty acids (NEFA) levels, decreased maternal IL‐6, TNFα and leptin levels compared to BFIII females (p < .05). BFIII sows were associated with significantly decreased newborn TG levels, increased newborn glucose, IL‐6 and TNFα levels compared to BFI or BFII sows (p < .05). BFI and BFII females had significantly decreased placental TG, NEFA and cholesterol (CHOL) contents compared to BFIII females (p < .05). Moreover, decreased CD36, FATP1, FABP4, and FABP1 mRNA and protein and FATP4 protein expression, and increased LPL activity were also observed in BFIII group compared with BFII group (p < .05). PPARγ mRNA and protein and lipogenic genes such as SREBP‐1c, ACSL1, ACCα, FAS and SCD mRNA expression were downregulated or upregulated, respectively, in the placentas of BFIII sows compared to BFI or BFII sows (p < .05). Overall, this study demonstrated that there is no advantage, in terms of litter live size, litter weight and placental FA transport and metabolism, in performing the mating of sows with BF>19 mm.     

Effects of Dietary Supplementation of β‐hydroxy‐β‐methylbutyrate on Sow Performance and mRNA Expression of Myogenic Markers in Skeletal Muscle of Neonatal Piglets

The effects of dietary β‐hydroxy‐β‐methylbutyrate (HMB) supplementation during gestation on reproductive performance of sows and the mRNA expression of myogenic markers in skeletal muscle of neonatal pigs were determined. At day 35 of gestation, a total of 20 sows (Landrace × Yorkshire, at third parity) were randomly assigned to two groups, with each group receiving either a basal diet or the same diet supplemented with 4 g/day β‐hydroxy‐β‐methylbutyrate calcium (HMB‐Ca) until parturition. At parturition, the total and live litter size were not markedly different between treatments, however, the sows fed HMB diet had a decreased rate of stillborn piglets compared with the sows fed the control (CON) diets (p < 0.05). In addition, piglets from the sows fed HMB diet tended to have an increased birth weight (p = 0.08), and a reduced rate of low birth weight piglets (p = 0.05) compared with piglets from the CON sows. Nevertheless, lower feed intake during lactation was observed in the sows fed the HMB diet compared with those on the CON diet (p < 0.01). The relative weights of the longissimus dorsi (LD) and semitendinosus (ST) muscle were higher (p < 0.05) in neonatal pigs from the HMB than the CON sows. Furthermore, maternal HMB treatment increased the mRNA levels of the myogenic genes, including muscle regulatory factor‐4 (MRF4, p < 0.05), myogenic differentiation factor (MyoD) and insulin‐like growth factor‐1 (IGF‐1, p < 0.01). In conclusion, dietary HMB supplementation to sows at 4 g/day from day 35 of gestation to term significantly improves pregnancy outcomes and increases the expression of myogenic genes in skeletal muscle of neonatal piglets, but reduces feed intake of sows during lactation.     

Methyl donors dietary supplementation to gestating sows diet improves the growth rate of offspring and is associating with changes in expression and DNA methylation of insulin‐like growth factor‐1 gene

The study aimed to investigate the effects of maternal dietary methyl donors on the performance of sows and their offspring, and the associated hepatic insulin‐like growth factor‐1 (IGF‐1) expression of the offspring. A total of 24 multiparous sows were randomly fed the control (CON) or the CON diet supplemented with methyl donors (MD) at 3 g/kg betaine, 15 mg/kg folic acid, 400 mg/kg choline and 150 μg/kg VB₁₂, from mating until delivery. After farrowing, sows were fed a common lactation diet through a 28‐days lactation period and six litters per treatment were selected to be fed until at approximately 110 kg BW. Maternal MD supplementation resulted in greater birthweight (p < 0.05) and increased the piglet weights (p < 0.01) and litter weights (p < 0.05) at the age of day 28, compared with that in CON group. The offspring pigs in the MD group had greater ADG (p < 0.05) and tended to lower F:G ratio (p = 0.07) compared with that of CON group from day 28 to 180 of age. The offspring pigs from MD group had greater serum IGF‐1 concentrations and expressions of hepatic IGF‐1 gene and muscular IGF‐1 receptor (IGF‐1r) protein at birth (p < 0.05), and greater hepatic IGF‐1 protein (p = 0.03) and muscular IGF‐1r gene expressions (p < 0.05) at slaughter, than that from the CON group. Moreover, the methylation at the promoter of IGF‐1 gene in the liver of newborn piglets and finishing pigs was greater in the MD group than that of the CON group (p < 0.05). In conclusion, maternal MD supplementation throughout gestation could enhance the birthweight and postnatal growth rate of offspring, associated with an increased expression of the IGF‐1 gene and IGF‐1r, as well as the altered DNA methylation of IGF‐1 gene promotor.     

Changes of leukocyte counts and expression of pro‐ and anti‐inflammatory cytokines in peripheral leukocytes in periparturient dairy cows with retained fetal membranes

In dairy cows, retained fetal membranes (RFM) affect reproductive performance. The aim of this study was to examine the leukocyte counts and the gene expression of tumour necrosis factor α (TNFα), interleukin 1β (IL‐1β), IL‐8, and IL‐10 in polymorphonuclear leukocytes (PMNs) and peripheral blood mononuclear cells (PBMCs) in cows with (n = 5) or without (n = 5) RFM during the peripartum period. The lymphocyte counts in RFM cows were higher than those in control cows throughout the experiment (p < .05). The expression of IL‐8 in PMNs of control cows was higher (p < .05) compared with that of RFM cows postpartum. In cows with RFM, IL‐1β expression was higher (p < .05) in PMNs at 6 weeks postpartum whereas the expression of IL‐1β was lower (p < .05) in PBMCs at 4 weeks postpartum. The expression of IL‐10 in PBMCs of control cows was higher (p < .05) than that of RFM cows at 2 weeks prepartum and 4 weeks postpartum. Taken together, our data indicate that changes of gene expression of pro‐ and anti‐inflammatory cytokines in RFM cows might be associated with the delayed placental separation and development of uterine inflammation in RFM cows.     

GnRH and prostaglandin‐based synchronization protocols as alternatives to progestogen‐based treatments in sheep

The study investigated, for cycling sheep, synchronizing protocols simultaneously to the standard “P” protocol using progestogens priming with intravaginal devices and gonadotropin. In November 2014, 90 adult Menz ewes were assigned to either the “P” protocol, “PGF” treatment where oestrus and ovulation were synchronized using two injections of prostaglandin 11 days apart or a “GnRH” treatment where the ewes had their oestrus and ovulation synchronized with GnRH (day 0)–prostaglandin (day 6)–GnRH (day 9) sequence. The ewes were naturally mated at the induced oestrus and the following 36 days. Plasma progesterone revealed that 92% of the ewes were ovulating before synchronization and all, except one, ovulated in response to the applied treatments. All “P” ewes exhibited oestrus during the 96‐hr period after the end of the treatments in comparison with only 79.3% and 73.3% for “PGF” and “GnRH” ewes, respectively (p < .05). Onset and duration of oestrus were affected by the hormonal treatment (p < .05); “GnRH” ewes showed oestrus earliest and had the shortest oestrous duration. Lambing rate from mating at the induced oestrus was lower for “P” than for “PGF” ewes (55.6% and 79.3%, respectively; p < .05). The same trait was also lower for “P” than for “PGF” and “GnRH” ewes (70.4%, 89.7% and 86.7%, respectively; p < .05) following the 36‐day mating period. Prostaglandin and GnRH analogue‐based protocols are promising alternatives for both controlled natural mating and fixed insemination of Menz sheep after the rainy season when most animals are spontaneously cycling.     

Effects of Inulin Supplementation in Low‐ or High‐Fat Diets on Reproductive Performance of Sows and Antioxidant Defence Capacity in Sows and Offspring

This experiment was conducted to investigate the effects of inulin supplementation in low‐ or high‐fat diets on both the reproductive performance of sow and the antioxidant defence capacity in sows and offspring. Sixty Landrace × Yorkshire sows were randomly allocated to four treatments with low‐fat diet (L), low‐fat diet containing 1.5% inulin (LI), high‐fat diet (H) and high‐fat diet containing 1.5% inulin (HI). Inulin‐rich diets lowered the within‐litter birth weight coefficient of variation (CV, p = 0.05) of piglets, increased the proportion of piglets weighing 1.0–1.5 kg at farrowing (p < 0.01), reduced the loss of body weight (BW) and backfat thickness (BF) during lactation (p < 0.05) and decreased the duration of farrowing as well as improved sow constipation (p < 0.05). Sows fed fat‐rich diets gained more BW during gestation (p < 0.01), farrowed a greater number of total (+1.65 pigs, p < 0.05) and alive (+1.52 pigs p < 0.05) piglets and had a heavier (+2.06 kg, p < 0.05) litter weight at birth as well as a decreased weaning‐to‐oestrous interval (WEI, p < 0.01) compared with sows fed low‐fat diets. However, it is worth noting that the H diet significantly decreased the serum activities of superoxide dismutase (T‐SOD) and glutathione peroxidase (GSH‐Px) and increased the serum malondialdehyde (MDA) levels in sows and piglets (p < 0.05). In contrast, HI diet enhanced the activities of T‐SOD and GSH‐Px and decreased the serum MDA concentrations (p < 0.05) in sows and piglets. In summary, the fat‐rich diets fed to sows during gestation had beneficial effects on reproductive performance, but aggravated the oxidative stress in sow and piglets. Inulin‐rich diets fed to sow during gestation had beneficial effects on within‐litter uniformity of piglet birthweight and enhanced the antioxidant defence capacity of sows and piglets.     

Maternal dietary linoleic acid supplementation promotes muscle fibre type transformation in suckling piglets

As meat quality is basically dependent on muscle fibre characteristics, it is important to know how muscle fibres are regulated and transformed. This study aimed to investigate the effect of maternal dietary supplementation on muscle fibre types using 3% saturated fatty acid (palmitic acid, PA) or 3% unsaturated fatty acid (linoleic acid, LA) from 80 days of gestation to the weaning of offspring (25 days post‐natal). The results indicated that higher mRNA levels of MyHCI type genes were found in the soleus muscles of piglets that suckled from LA‐supplemented sows than from PA‐supplemented sows. In addition, LA treatment increased the gene expression of the type I muscle fibre marker troponin I (p < 0.01), suggesting that LA promoted muscle fibre type transformation to type I fibres. Moreover, PGC‐1α (p < 0.01) and MEF2c (p < 0.05) mRNA levels were higher in the piglets from the LA treatment group than in those from the PA treatment group. Furthermore, LA supplementation also significantly increased AMP‐activated protein kinase (AMPK) mRNA levels (p < 0.05), which is an upstream regulator of PGC‐1α. Collectively, these findings demonstrated that maternal dietary LA supplementation promoted muscle fibre transformation to type I fibre and that this process may be mediated through an AMPK‐dependent pathway.     

Effects of Saccharomyces cerevisiae supplementation on growth performance,plasma metabolites and hormones,and rumen fermentation in Holstein calves during pre‐ and post‐weaning periods

This study aimed to evaluate the effects of supplementing Saccharomyces cerevisiae (SC) during the pre‐ and post‐weaning periods on growth, metabolic and hormonal responses, and rumen fermentation in calves. Three‐week‐old Holstein calves were assigned to either control (n = 12) or SC group (n = 12), the latter of which received 2 × 10⁹ cfu/day of SC. The experiment was conducted over a period of 7 weeks around weaning. Daily gain (DG) in the SC group was higher (p < .05) than that in the control group. In the SC group, plasma glucose, insulin, and growth hormone (GH) concentrations were higher (p < .05) and concentrations of glucagon and insulin‐like growth factor 1 (IGF‐1) tended to be higher (p < .1) than in the control group. Proportion of rumen propionate and concentration of rumen ammonia nitrogen at 10 weeks of age were greater (p < .05) in the SC group than that in the control group. Supplementation of SC around weaning may improve dietary nutrient and energy availability and increase plasma GH and IGF‐1 concentrations. These changes observed in SC‐supplemented calves could be closely related to the improvement of DG.     

Lactobacillus rhamnosus GG promotes M1 polarization in murine bone marrow‐derived macrophages by activating TLR2/MyD88/MAPK signaling pathway

Lactobacillus rhamnosus GG (LGG) is increasingly applied in functional food products and acts as a probiotic model in nutritious and clinical studies. Increasing evidences have revealed the immune modulation of LGG on macrophages. The aim of this study is to investigate the effect of LGG on macrophage polarization of murine bone marrow‐derived macrophages (BMDMs). BMDMs were treated with 10⁸ colony‐forming units (CFU)/ml LGG for 1.5, 3, and 6 hr. Results showed that LGG obviously upregulated the mRNA expression of M1‐associated cytokines (p < .05), including interleukin‐1 beta (IL‐1β), IL‐6, tumor necrosis factor‐alpha (TNF‐α), and inducible nitric oxide synthase (iNOS), whereas had no effect on the expression of M2‐associated markers (p > .05), including arginase 1 (Arg1), mannose receptor, and chitinase‐like protein 3 (YM1). Furthermore, LGG markedly increased the expression of pro‐inflammatory cytokines (IL‐12p40, cyclooxygenase‐2 [COX‐2], and interferon‐γ [IFN‐γ]) (p < .05) and anti‐inflammatory cytokines (IL‐10, IL‐4, and transforming growth factor‐β [TGF‐β]) (p < .05). In addition, we also found that TLR2/MyD88/MAPK signaling pathway was required for LGG‐induced M1 macrophage polarization and M1‐related cytokines expression. Together, these findings demonstrate that probiotic LGG facilitates M1 polarization of BMDMs, suggesting that LGG may have an immunotherapeutic potential in regulating the host defense against pathogen invasion.     

Investigation of carryover effect of prior fibre consumption on growth,serum and tissue metabolic markers in Ossabaw pigs fed a high‐fat diet

Carryover effect of prior fibre consumption on metabolic markers was investigated. Treatments were arranged in 2 × 2 factorial with 2 fibre sources, 4% inulin or cellulose (Solka‐Floc®) and fat levels (5 or 15%) for the low‐fat diet (LFD) and high‐fat diet (HFD) respectively. Pigs were fed the two fibre diets for the first 56d (nursery phase), and thereafter fed either the LFD or HFD containing no added fibre source from d56 to 140 (growing phase). Pigs on the HFD were heavier (p = .05) than those on LF (64.61 vs. 68.38 kg), regardless of prior fibre type consumed. Pigs that were fed cellulose during the nursery and later fed the HFD had the highest ADG (p < .05). Feeding the HFD resulted in higher back fat (BF) (13.41 and 18.18 ± 0.12 mm for LFD and HFD, respectively; p < .01). The HFD resulted in higher (p < .01) insulin (0.014 and 0.016 ± 0.001 mg/L for LF and HF respectively) and glucose (100.89 and 125.03 ± 4.39 mg/dl for LF and HF respectively) concentrations in the serum. Inulin increased (p ≤ .02) jejunal expression of SREBP‐1c and CL‐4, but reduced (p < .05) TNFɑ and IL‐6 expression in the ileum. Alpha‐diversity was significantly different (p < .05) between the inulin and cellulose fed pigs at the end of the nursery and finishing phases. Therefore, inulin feeding before a HFD may lead to reduction in ADG and inflammatory markers in the small intestine of pigs, and thus prevent future metabolic disorders.     

Genetic differences in oestrous signs and oestrogen metabolism‐related genes between Chinese Mi and European Landrace‐Large White pigs

Oestrous signs affect timely mating and reproductive efficiency in swine breeding herds. To study the genetic difference of oestrous signs between Chinese and European pigs, 100 Landrace‐Large White (LLW) cross gilts and 50 Chinese Mi gilts were assessed for oestrous signs and the concentrations of serum estradiol‐17β and progesterone were determined. The genotype of 39 single nucleotide polymorphisms (SNPs) in 11 oestrogen metabolism and function‐related genes was determined by Sequenom iPLEX platform. Compared with LLW gilts, Mi gilts had longer time of standing reflex (p < .001), higher scores of vulva reddening (p = .001) and greater serum estradiol‐17β concentration (p < .01). Gilts with greater serum estradiol‐17β concentrations also had greater (p < .05) scores for oestrous signs. Genetic polymorphisms of nine genes in oestrogen metabolism pathways had significant differences (p < .05) between LLW and Mi gilts. There were three and six haploblocks of SNPs in LLW and Mi, respectively. Compared with LLW, the distribution of haplotypes was more centralized in Mi pigs. Genetic polymorphisms of oestrogen metabolism‐related genes have considerable differences between Chinese Mi and European LLW pigs. Because of the important roles of oestrogen during the oestrus, some genes of oestrogen metabolism pathway could be considered as candidate genes for oestrous signs.     

Effectiveness of controlled internal drug release device treatment to alleviate reproductive seasonality in anestrus lactating or dry Barki and Rahmani ewes during non‐breeding season

This study aimed to evaluate the effectiveness of hormonal treatments on ovarian activity and reproductive performance in Barki and Rahmani ewes during non‐breeding season. Forty‐eight multiparous ewes, 24 Barki and 24 Rahmani ewes were divided into two groups, 12 lactating and 12 dry ewes for each breed. Controlled internal drug release (CIDR) device was inserted in all ewes for 14 days in conjunction with intramuscular 500 IU equine chronic gonadotrophin (eCG) at day of CIDR removal. Data were analysed using PROC MIXED of SAS for repeated measures. Breed, physiological status and days were used as fixed effects and individual ewes as random effects. Barki ewes recorded higher (p < .05) total number of follicles, number of large follicles, serum estradiol concentration and estradiol: progesterone (E₂:P₄) ratio compared to Rahmani ewes. Lactating ewes recorded higher (p < .05) number of small follicles and lower concentration of total antioxidant capacity (TAC) compared to dry ewes. Number and diameter of large follicles recorded the highest (p < .05) values accompanied with disappearance of corpora lutea at day of mating. Serum progesterone concentration recorded lower (p < .05) value at day of mating and the highest (p < .05) value at day 35 after mating. CIDR‐eCG protocol induced 100% oestrous behaviour in both breeds, but Rahmani ewes recorded longer (p < .05) oestrous duration compared to Barki. Conception failure was higher (p < .05) in Barki compared to Rahmani ewes. In conclusion, CIDR‐eCG protocol was more potent in improving ovarian activity in Barki compared to Rahmani ewes, but this protocol seems to induce hormonal imbalance in Barki ewes that resulted in increasing conception failure compared to Rahmani ewes.     

Effects of L‐citrulline supplementation on heat stress physiology,lactation performance and subsequent reproductive performance of sows in summer

Lactating sows are susceptible to heat stress (HS). Part of the thermoregulatory response to HS is to increase peripheral blood flow, which is mediated in part by the vasodilator, nitric oxide (NO). Therefore, the aim of this experiment was to determine the effect of supplementation of L‐citrulline, a NO precursor, on symptoms of HS, lactation performance and subsequent reproductive performance of sows in summer. A total of 221 summer farrowing mixed parity sows were fed either a control diet or supplemented with 1% L‐citrulline upon entry to the farrowing house (6 ± 1.8 days for mean ± standard deviation [SD] before farrowing) until weaning (26 ± 1.5 days). The average daily minimum and maximum temperature in the farrowing house was 21.0 ± 1.88 and 29.2 ± 3.82°C (mean ± SD). Rectal temperature, respiration rate, and plasma and urinary nitrite and nitrate (NOx) of sows were measured on the 19th day post‐farrowing. Supplemental L‐citrulline in the diet did not affect the number of piglets born alive, feed intake of sows, body weight or backfat thickness of sows at weaning, or litter weight gain. L‐citrulline tended to reduce piglet pre‐weaning mortality rate from 18.6% to 15.6% (p = 0.058). L‐citrulline reduced the respiration rate of sows compared to the control diet at 17:00 hr (Time × Diet, p < 0.001); however, rectal temperature was not affected. L‐citrulline tended to increase urinary NOx concentrations (127 vs. 224 µM, p = 0.057) but not plasma NOx concentrations. L‐citrulline did not affect farrowing rate or number of piglets born alive in the subsequent parity. In conclusion, L‐citrulline supplementation reduced respiration rate of lactating sows and reduced piglet pre‐weaning mortality rate in summer. Whether the effects were due to a NO‐dependent mechanism requires further validation.     

Ovine sperm DNA oxidation quantification using an 8‐OHdG immunodetection assay

Our aim was to optimize 8‐hydroxy‐2′‐deoxyguanosine (8‐OHdG) immunodetection in order to detect DNA damage caused by oxidative stress that may not be detected by other DNA integrity analysis techniques, especially due to the high compaction of DNA in ruminants. Semen samples from 6 rams were cryopreserved. After thawing, samples were subjected to the DNA oxidation quantification using an 8‐OHdG immunodetection assay by flow cytometry. We have evaluated two different incubation times (30 min vs. overnight) at 4°C of the primary antibody (monoclonal anti‐8‐OHdG antibody). We have also compared the results of this technique with the sperm chromatin structure assay (SCSA^®). The analysis revealed that there were no significant differences (p > .05) between different incubation times. However, overnight incubation seems to cause more non‐specific binding of the secondary antibody. Significant differences (p < .05) between subjects and oxidation controls (8 M H₂O₂/800 μM FeSO₄?7H₂O) were evident. We can conclude that the 8‐OHdG immunodetection assay for DNA oxidation quantification of ram sperm can be performed subjecting sperm samples to a very high oxidative treatment.     

The effect of select seminal plasma proteins on endometrial mRNA cytokine expression in mares susceptible to persistent mating‐induced endometritis

In the horse, breeding induces a transient endometrial inflammation. A subset of mares are unable to resolve this inflammation, and they are considered susceptible to persistent mating‐induced endometritis PMIE Select seminal plasma proteins cysteine‐rich secretory protein‐3 (CRISP‐3) and lactoferrin have been shown to affect the innate immune response to sperm in vitro. The objective of this study was to determine whether the addition of CRISP‐3 and lactoferrin at the time of insemination had an effect on the mRNA expression of endometrial cytokines in susceptible mares after breeding. Six mares classified as susceptible to PMIE were inseminated during four consecutive oestrous cycles with treatments in randomized order of: 1 mg/ml CRISP‐3, 150 μg/ml lactoferrin, seminal plasma (positive control) or lactated Ringer's solution (LRS; negative control) to a total volume of 10 ml combined with 1 × 10⁹ spermatozoa pooled from two stallions. Six hours after treatment, an endometrial biopsy was obtained for qPCR analysis of selected genes associated with inflammation (pro‐inflammatory cytokines interleukin (IL)‐1β, IL‐8, tumour necrosis factor (TNF)‐α, interferon (INF)‐γ, anti‐inflammatory cytokines IL‐1RN and IL‐10, and inflammatory‐modulating cytokine IL‐6). Seminal plasma treatment increased the mRNA expression of IL‐1β (p = .019) and IL‐8 (p = .0068), while suppressing the mRNA expression of TNF (p = .0013). Lactoferrin also suppressed the mRNA expression of TNF (p = .0013). In conclusion, exogenous lactoferrin may be considered as one modulator of the complex series of events resulting in the poorly regulated pro‐inflammatory response seen in susceptible mares.     

Differential gene expression of Eph‐ephrin A1 and LEPR‐LEP with high or low number of embryos in pigs during implantation

The objective of this study was to ascertain whether mRNA and protein expressions of implantation‐related genes (erythropoietin‐producing hepatocellular receptor–ligand A1, Eph‐ephrin A1 and leptin receptor–leptin, LEPR‐LEP) differed between pigs with high and low number of embryos, and whether these differences in gene expression might affect embryo implantation. Experimental pig groups (n = 24) for high and low number of embryos were prepared by altering the number of eggs ovulated in pre‐pubertal gilts treated with 1.5 × (High) or 1.0 × (Low) PG600 ([400 IU PMSG + 200 IU hCG]/dose, AKZO‐NOBEL). Gilts expressing oestrus were artificially inseminated twice and maintained in breeding and gestation until the reproductive tract was collected on day 22 of pregnancy. At slaughter, the reproductive tracts from each pregnant gilt from each treatment were immediately processed to collect samples for RNA and protein analysis. Within each gilt, three conceptus points were sampled, one from each horn and then a random conceptus within the tract. At each conceptus point, endometrial attachment site, chorion–allantois and embryo were collected and immediately frozen in liquid nitrogen. Number of corpus luteum (CL) (35.4 vs. 12.6) and total embryo number (18.8 vs. 10.2) were greater in the high‐embryo compared to the low‐embryo group, respectively (p < .05). Real‐time qPCR results showed that Eph‐ephrin A1 mRNA expression was less in the high‐embryo (p < .05) compared to the low‐embryo group. In addition, Western blotting analysis indicated that Eph‐ephrin A1 and LEP protein expression at endometrial attachment site in high‐embryo was less (p < .05) compared to low‐embryo group. It was also noted that mRNA expression of Eph‐ephrin A1 and LEPR‐LEP was greater in pregnant than non‐pregnant gilts (p < .05). Moreover, mRNA expression of Eph‐ephrin A1 (p < .05) and LEPR‐LEP was greatest at endometrial attachment site among all three tissues. There was a positive correlation between expressions of Eph‐ephrin A1, LEPR‐LEP and embryo length with the correlation coefficient 0.31–0.59. For Eph‐ephrin A1, the highest correlation coefficient appeared between Eph A1 expression and normal embryo number, between ephrin A1 expression and embryo length. For LEPR‐LEP, the highest correlation coefficient appeared between LEPR‐LEP expression and ovary weight (0.79 for both, p < .05), followed by embryo length and weight. The results of this study suggest that low expression of Eph‐ephrin A1 and LEPR‐LEP is somehow related to increased embryo number during implantation and that endometrial attachment site might be the main target tissue of these gene products. Yet, the increased expression of Eph‐ephrin A1 and LEPR‐LEP appeared associated with increased embryo growth (length and weight) and ovary weight, Eph‐ephrin A1 and LEPR‐LEP might play roles in the regulation of embryo implantation in pigs.     

Effect of dietary supplementation of calcium butyrate on growth performance,carcass traits,intestinal health and pro‐inflammatory cytokines in Japanese quails

The objective of the present study was to evaluate the potential effect of dietary calcium butyrate on growth performance, carcass traits and gut health in Japanese quails. In total, 320 one‐day‐old Japanese quails were randomly assigned to 4 equal treatments, with 8 replicates of 10 Japanese quails, for 4 weeks. The Japanese quails in control treatment were fed control diet whereas in the other treatments the Japanese quails were fed diet supplemented with calcium butyrate at 0.3, 0.5 and 0.7 g/kg diet. Data concerning performance measurements were recorded weekly. In addition, eight Japanese quails (one/replicate) from each treatment were selected randomly for serum collection to measure pro‐ and anti‐inflammatory cytokines. Pooled faecal samples from each replicate of each treatment were also collected at three time points (0, 2 and 4 weeks) for count E. coli and C. perfringens. The results showed that after 7 days of the experimental period, Japanese quails fed calcium butyrate supplemented diet at 0.7 g/kg showed a greater (p < .05) body weight and a favourable (p < .05) feed conversion ratio than the other treatments. Moreover, serum superoxide dismutase and catalase activities were increased (p < .05) in Japanese quails fed calcium butyrate supplemented diet at 0.7 g/kg. Calcium butyrate supplementation at 0.7 g/kg was associated with reduction (p < .05) in TNF‐α, IL‐6 and IL1‐β, while IL‐10 was increased (p < .05). In addition, after 2 weeks of calcium butyrate supplementation, a reduction (p < .05) in E. coli and C. perfringens counts was observed in excreta of Japanese quails fed 0.5 and 0.7 g calcium butyrate/kg diets. It is concluded that calcium butyrate supplementation improves body weight gain, reduces E. coli and C. perfringens counts and has anti‐inflammatory/anti‐oxidant effect in Japanese quails.     

Effect of dietary threonine on laying performance and intestinal immunity of laying hens fed low‐crude‐protein diets during the peak production period

Threonine (Thr) may be a limiting amino acid for laying hens fed diets with lowered protein level. An experiment was conducted to examine laying performance, and the intestinal immune function of laying hens provided diets varying in digestible Thr levels. Lohmann Brown laying hens (n = 480), 28 weeks of age, were allocated to six dietary treatments, each of which included five replicates of 16 hens. Dietary crude protein (CP) 16.18% diet was offered as the positive control diet. L‐Thr was added to the negative diet (14.16% CP) by 0, 1.0, 2.0, 3.0 and 4.0 g/kg, corresponding 0.44%, 0.43%, 0.49%, 0.57%, 0.66% and 0.74% digestible Thr. At 40 weeks, a reduction in CP level decreased laying performance (p < 0.05). In the low CP, increasing dietary Thr increased (p < 0.05) egg production and egg mass and rose to a plateau between 0.57% and 0.66%. The hens fed 0.66% Thr showed the lowest value (p < 0.05) of feed conversion ratio (FCR). Serum level of uric acid showed the lowest values (p < 0.05) at 0.57–0.66%. In addition, serum‐free Thr maximized (p < 0.05) between 0.66% and 0.74%. Digestive trypsin activity decreased (p < 0.05) when hens fed the low‐CP diet compared with hens fed CP (16.18%) and hens fed 0.57–0.66%. Expressions of ileal MUC2 mRNA maximized (p < 0.05) at 0.66% Thr. Occludin mRNA increased with increasing Thr level (p < 0.05). sIgA mRNA reached to the maximum level (p < 0.05) at 0.66% and 0.74% Thr. INF‐γ mRNA reached to the lowest level (p < 0.05) at 0.65%. Expressions of ileal IL‐2, IL‐6, IL‐1β mRNA decreased with increasing Thr level (p < 0.05). In conclusion, Thr supplementation resulting in optimal laying performance and stimulated the mucosal immune system, suggesting that it is a limiting amino acid in the low‐crude‐protein diet of laying hens during the peak production period.     

Changes in in vitro ruminal and post‐ruminal degradation of tropical tannin‐rich legumes due to varying levels of polyethylene glycol

We evaluated the effects of tannins from Flemingia macrophylla (CIAT 17403) and Calliandra calothyrsus (San Ramón CIAT 22310 and Patulul CIAT 22316) on in vitro ruminal and post‐ruminal dry matter and apparent protein degradation. For each tannin source (legumes), different dosages of polyethylene glycol (PEG) (8000 Da) in McDougall buffer were added to achieve ratios of 0:3, 1:3, 2:3 and 3:3 PEG:condensed tannin (CT). Ruminal fluid mixed with McDougall buffer (1:4) was added to tubes containing only legume foliage (control) or PEG‐treated legume foliage. For both Calliandra varieties, a higher ruminal dry matter degradation was observed at a PEG:CT ratio of 3:3. For F. macrophylla, no differences were found between 2:3 and 3:3 ratios (p > 0.05), indicating that a PEG:CT ratio of 2:3 might be enough to bind tannins. Increasing PEG:CT ratios increased apparent ruminal degraded protein and ammonia concentration (p < 0.0001) differing among species (species × ratio: p < 0.0001). The degradation of bypass crude protein (dBCP) was influenced by both legume type and PEG:CT ratio (p < 0.0001). For Patulul, as PEG:CT ratio increased, dBCP increased, but after tannin ratio of 2:3, there was not a significant increase, and for San Ramón, dBCP degradation was higher as PEG:CT ratio increased up to 2:3. For Flemingia, dBCP was higher than PEG:CT ratio of 0:3 but not different among 1:3, 2:3 or 3:3. Low concentration of CT (116 mg/g DM) increased the proportion of protein digested in the abomasum, but higher levels of CT (252 mg/g) clearly reduced the proportion of digested CP. For Flemingia, PEG:CT ratio of 2:3 is enough to inactivate tannins, while PEG:CT ratio of 3:3 was needed for Calliandra and consequently increased ruminal degradation of dry mater (rdDM), and crude protein (rdCP), total degradation of dry matter (tdDM), crude protein (tdCP) and ammonia levels.     

The effect of Epigallocatechin‐3‐gallate on small intestinal morphology,antioxidant capacity and anti‐inflammatory effect in heat‐stressed broilers

This study was to investigate the effects of Epigallocatechin‐3‐gallate (EGCG) on intestinal morphology, antioxidant capacity and anti‐inflammatory response in heat‐stressed broiler. A total of 192 2‐week‐old Arbour Acres broilers chickens were divided into four groups with six replicates per group and eight chickens per replicate: one thermoneutral control group (28°C, group TN), which was fed the basal diet; and three cyclic high‐temperature groups (35°C from 7:00 to 19:00 hr; 28°C from 19:00 hr to 7:00 hr, heat stress group), which were fed the basal diet supplementation with EGCG 0 mg/kg (group HS0), 300 mg/kg (group HS300) and 600 mg/kg (group HS600). The gut morphology and intestinal mucosal oxidative stress indicators, as well as intestinal barrier‐related gene expression, were analysed. The results showed that compared with group TN, heat stress reduced the villus height (VH), activities of glutathione peroxidase (GSH‐Px), superoxide dismutase (SOD)and catalase (CAT), increased the crypt depth (CD) and malondialdehyde (MDA)content at 21, 28 and 35 days (p < 0.05). After the heat‐stressed broilers were supplemented with EGCG, VH, VH/CD (V/C), and the activities of GSH‐Px, SOD and CAT were increased, and CD and MDA content were reduced compared with those in group HS0 without EGCG supplementation at 21, 28 and 35 days (p < 0.05). The EGCG supplementation promoted the gene expression of nuclear factor‐erythroid 2‐related factor 2 (Nrf2), Claudin‐1, Mucin 2 (Muc2) and alleviated the nuclear factor‐kappa B (NF‐κB) and lipopolysaccharide‐induced tumour necrosis factor (LITAF) gene expression compared with group HS0 (p < 0.05). Moreover, intestinal morphology was strongly correlated with antioxidant ability and inflammatory response. In conclusion, EGCG alleviated the gut oxidative injury of heat‐stressed broilers by enhancing antioxidant capacity and inhibiting inflammatory response.